During pregnancy, a variety of cell types of fetal origin cross the placenta and circulate within maternal peripheral blood. Fetal cells have been detected in maternal circulation during certain stages of gestation (Holzgreve et al., 1992, J. Reprod. Med., 37:410; Simpson, et al., 1993, JAMA, 270:2357). Four fetal cell types in maternal blood that are accessible to prenatal diagnosis are lymphocytes, trophoblasts, stem cells and nucleated red blood cells. These cells provide a potential source of information about the gender and genetic makeup of the developing fetus. One particular fetal cell type within maternal blood that has been demonstrated to be useful for analyzing fetal DNA is the nucleated erythrocyte. The feasability of using fetal cells from the maternal circulation for diagnostic purposes, however, is greatly hindered by the fact that fetal cells are present in maternal blood in only very limited numbers. In addition, most fetal cells (with the exception of trophoblasts) cannot be distinguished from maternal cells on the basis of morphology alone; instead, identification must be based upon detection of either fetal cell markers or fetal DNA.
Detection of fetal cells in maternal blood can be improved by enrichment for fetal cells within the mixture of fetal and maternal cells and/or by separation of fetal cells from maternal cells. One approach that has been used to achieve enrichment for and separation of fetal cells within a maternal blood sample utilizes antibodies (Abs) specific for a particular fetal cell type to couple to and capture fetal cells or to label fetal cells. For example, as described in U.S. Pat. No. 5,641,628, fetal-specific, detectably labeled antibodies are used to label fetal cells and, when bound to these fetal cells, facilitate separation of these cells from maternal components by flow cytometry.
Another method of separating target cells from heterogenous cell populations in bodily fluids, such as blood, has been through the use of beds of particles which carry sequestering agents that are selected to capture a specific cell; examples of such sequestering agents are antibodies (Abs) that are directed at a ligand carried on the exterior surface of the target cells. The bodily fluid may be caused to flow through a stationary bed of such particles, or a group or bed of such particles may be caused to move, as by gravity, through a sample of the bodily fluid in question. Oftentimes this separation using such beads has taken place in a vertical column, and it is sometimes referred to as the “column” separation method. S. M. Gomez et al. in Biotechnol. Prog. 1999, 15, 238-244, use macroscopic polystyrene beads (100-170 microns) carrying CD45-Abs to separate a minor subpopulation of target cells from a large population of “bystander” cells.
There have been at least attempts to separate a fairly wide variety of cells by employing beads of this general type; particularly, beads have been used to separate fetal cells from maternal cells present in maternal blood of a pregnant woman. Such a method of separating target cells from a heterogenous population of cells suspended in a liquid medium is disclosed, for example, in International Application WO 94/26104. Moreover, U.S. Pat. No. 5,766,843 teaches the bonding of anti-CD45 antibodies to the exterior surface of solid supports, such as magnetic beads, which are then used to selectively bind to white blood cells. This patent also indicates that such beads coated with streptavidin are commercially available from Calbiochem and that antibodies to surface antigens on a wide variety of target cells may readily be attached thereto. U.S. Published Patent Application No. 2004/0018509 mentions the use of commercially available “Dynabeads” having magnetic cores, which are coated with antibodies, for removing placenta-derived trophoblast cells in the blood of pregnant women. Published Application 2003/0153028 mentions a number of types of polymeric beads that may be used as substrates to capture target cells of interest or to be removed from a blood sample.
U.S. Pat. No. 4,836,928 demonstrates the use of a separation device wherein the interior lumens of hollow fibers of an appropriate porosity are coated with a hydrogel made from polyvinyl alcohol that is mixed with a liposome dispersion and polymerized. It is said that the liposomes then capture B-cells as a bodily fluid is caused to flow through the hollow fibers; the B-cells are subsequently separately recovered by treatment with a buffer that releases them.
U.S. Published Patent Application 2003/0229,393 discloses the use of a medical device, such as a stent, which is coated with a bio-compatible hydrogel matrix that contains antibodies which capture progenitor endothelial cells for the purpose of forming an endothelium on the surface of the medical device.
U.S. Published Patent Application 2004/0018509, teaches obtaining fetal DNA by first depleting a blood sample of the maternal cells prior to fetal cell isolation and sorting. In order to enrich the eventual proportion of fetal cells present, the maternal cells are selectively removed by incubating the cells with antibodies attached to a solid support, which antibodies bind antigens present on the cell surface of mature leukocytes.
Published U.S. Application 2004/0069710 discusses various types of bioseparation-type devices, including some that have used adsorbent particles that bind target compounds which are moved through filters. It advocates large scale processing through a hybrid bioseparation apparatus that uses adsorbent particles, such as chromatographic matrix beads, to sequester the bioproducts after they pass through a membrane that is coated with a polymeric coating that is essentially impermeable to water, and to the biomolecules to be isolated.
From the number of different vendors who are marketing beads useful for this purpose, it is quite clear that there is very substantial interest in achieving the separation of biomolecules by the so-called column method. Accordingly, the search has gone on for improved solid supports, such as beads, that can be employed to selectively sequester biomolecules of interest.